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Shelterin-Mediated Telomere Protection

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Shelterin-Mediated Telomere Protection GE52CH11_de_Lange ARI 26 October 2018 11:22 Annual Review of Genetics Shelterin-Mediated Telomere Protection Titia de Lange Laboratory of Cell Biology and Genetics, Rockefeller University, New York, NY 10065, USA; email: [email protected] Annu. Rev. Genet. 2018. 52:223–47 Keywords First published as a Review in Advance on telomere, shelterin, t-loop, DNA damage signaling, DNA repair, DNA September 12, 2018 replication The Annual Review of Genetics is online at genet.annualreviews.org Abstract https://doi.org/10.1146/annurev-genet-032918- For more than a decade, it has been known that mammalian cells use shelterin 021921 to protect chromosome ends. Much progress has been made on the mech- Copyright c 2018 by Annual Reviews. anism by which shelterin prevents telomeres from inadvertently activating All rights reserved DNA damage signaling and double-strand break (DSB) repair pathways. Shelterin averts activation of three DNA damage response enzymes [the ataxia-telangiectasia-mutated (ATM) and ataxia telangiectasia and Rad3- Access provided by Rockefeller University on 06/07/19. For personal use only. Annu. Rev. Genet. 2018.52:223-247. Downloaded from www.annualreviews.org related (ATR) kinases and poly(ADP-ribose) polymerase 1 (PARP1)], blocks three DSB repair pathways [classical nonhomologous end joining (c-NHEJ), alternative (alt)-NHEJ, and homology-directed repair (HDR)], and prevents hyper-resection at telomeres. For several of these functions, mechanistic in- sights have emerged. In addition, much has been learned about how shelterin maintains the telomeric 3 overhang, forms and protects the t-loop struc- ture, and promotes replication through telomeres. These studies revealed that shelterin is compartmentalized, with individual subunits dedicated to distinct aspects of the end-protection problem. This review focuses on the current knowledge of shelterin-mediated telomere protection, highlights differences between human and mouse shelterin, and discusses some of the questions that remain. 223 GE52CH11_de_Lange ARI 26 October 2018 11:22 INTRODUCTION Initial work on the molecular aspects of telomere function emphasized the end-replication prob- DDR: DNA damage lem, which originates from the inability of the canonical DNA replication pathway to complete response the duplication of the ends of linear DNA. Like most eukaryotes, mammals solve this problem using telomerase, which extends the 3 ends of chromosomes with telomeric DNA (16). Compared to the straightforward end-replication problem, the telomere end-protection problem is complicated because it arises from the myriad of cellular pathways that can sense and act on DNA ends. Early work by McClintock (94, 95) pointed to special features of telomeres that allowed them to escape the fusion reaction observed with broken chromosomes—the first hint that telomeres could avert nonhomologous end joining (NHEJ) and perhaps other double-strand break (DSB) repair pathways. In addition to inappropriate DNA repair reactions, telomeres need to pre- vent the activation of pathways that sense DNA damage. The early literature on eukaryotic DNA damage signaling pathways and checkpoints (reviewed in 15) did not address the obvious dilemma that cells need to distinguish DNA breaks from chromosome ends. Recent data indicate that telo- meres are threatened by at least seven distinct DNA damage response (DDR) pathways (Figure 1). The system that handles the many tasks related to the end-protection problem employs multiple mechanisms. Different threats to telomere integrity are dealt with using vastly different molecular strategies, each primarily targeting the initiation step of the DNA signaling or repair reaction. Remarkably, the various tricks to block the DDR are all performed by a single telomeric protein complex, shelterin (Figure 1). For some pathways, shelterin uses its own biochemical features to block the threat, and for others, it has co-opted proteins involved in genome maintenance. Although work in the past decade has brought many insights, we are far from understanding how this fascinating complex multitasks at telomeres. This review focuses on how shelterin solves the end-protection problem. The role of shelterin in recruiting and regulating telomerase and the nontelomeric functions of some of the shelterin subunits are discussed elsewhere (61, 91). SHELTERIN STRUCTURE AND DNA BINDING FEATURES Shelterin has evolved to bind specifically to the sequence and structure of mammalian telomeres (Figure 1a). Mammalian telomeres contain many kilobase pairs (kb) of tandem double-stranded (ds) TTAGGG repeats terminating in a 50–400 nucleotide 3 protrusion of single-stranded (ss) re- peats of the G-rich strand. Shelterin interacts with both ds and ss telomeric DNA and is sufficiently abundant to bind all telomeric DNA (Figure 1a–c) (128). Access provided by Rockefeller University on 06/07/19. For personal use only. Annu. Rev. Genet. 2018.52:223-247. Downloaded from www.annualreviews.org Human shelterin consists of six distinct proteins, TRF1, TRF2, Rap1, TIN2, TPP1, and a single version of POT1 (Figure 1a,d) (32). In contrast, rodent shelterin contains two closely related POT1 proteins, POT1a and POT1b, which arose through gene duplication (62, 142). The main interaction interfaces of the shelterin subunits have been established based on co- immunoprecipitation experiments, yeast two-hybrid analysis, and structural studies (Figure 1d). This work revealed that TRF1 and TRF2 bind to TIN2 using distinct interaction surfaces. TIN2 also binds to TPP1, which in turn binds to POT1 (or POT1a and POT1b in the mouse). Rap1 binds TRF2, completing the six-subunit complex. The binding interfaces between shelterin subunits are varied in structure and probably do not involve posttranslational modifications, despite the numerous reported modifications of the subunits (Figure 1d). The formation of shelterin also does not require interactions with DNA (44). The high-affinity DNA binding domains within shelterin are well established (Figure 1d). The Myb/SANT domains of TRF1 and TRF2 bind to duplex telomeric DNA with nanomolar affinity 224 de Lange GE52CH11_de_Lange ARI 26 October 2018 11:22 (44). High-affinity binding by TRF1 and TRF2 is dependent on the formation of homodimers, mediated by the TRFH domain (11, 12). Together, the TRF1 and TRF2 homodimers contribute four ds 5-TAGGGTT-3 recognition modules to shelterin (Figure 1a) (12, 99). Shelterin contains an additional Myb-like domain centrally located in Rap1. However, the surface charge of this Myb fold is unsuitable for binding to DNA (60), explaining its low affinity for DNA (1) and suggesting a 3’ GGATTGGGATT b GATTG GGG TPP1 ATT Shelterin GG POT1 G TRF1 TRF2 3' overhang TIN2 c Rap1p1 5' GGTT 3' 5' d TIN2 Tankyrase 1 FBOX4 BLM R13 115 F142 dsDNA TRF1 Alt- Acidic TRFH/dimerization Spl NLS Myb 439 aa (TERF1) 58 263 296 315 386 429 Rtel1 Apollo/SLX4 Rap1 TIN2 dsDNA TRF2 I121 F162 Ext Basic TRFH/dimerization RBM NLS iDDR Myb 542 aa (TERF2) 4284 286 317 358 396–409 448–473 489 538 DNA wrapping Branched DNA TRF2 S/TQ Rap1 BRCT Myb Acidic RCT (TERF2IP) 399 aa Phosphoserine 78101 128188 214 304 306 383 Ubiquitylation Shelterin core TRF2 TPP1 TRF1 Sumoylation TIN2 Muts T2 TRFH Arg methylation (TINF2) TPP1 T1 DC 355 aa (long form 451 aa) 200 250–265280–300 POT1 TIN2 TPP1 TEL patch OB POT1 TIN2 (ACD) Ser rich 544 aa 87244 266 320 354442 510 TPP1 POT1 OB1 OB2 OB3 (split) HJRL OB3 (split) 634 aa Access provided by Rockefeller University on 06/07/19. For personal use only. Annu. Rev. Genet. 2018.52:223-247. Downloaded from www.annualreviews.org 9141 161 278320 393 538 ssDNA e DDR pathway Shelterin subunit(s) Mechanism General repressor ATM kinase TRF2 (TIN2) t-loop None ATR kinase POT1a (POT1b) RPA exclusion None PARP1 TRF2, TIN2 Branched-DNA binding, ? Ku70/80 c-NHEJ TRF2 t-loop (iDDR, Rap1) CYREN (S/G2) alt-NHEJ TRF2 (TIN2, POT1a/b) t-loop, PARP1 repression, ? Ku70/80 HDR POT1a or POT1b + Rap1 ?, ? Ku70/80 Hyper-resection POT1a/b, TRF2 Repression of ATM/ATR 53BP1/Rif1/Rev7 (Caption appears on following page) www.annualreviews.org • Shelterin-Mediated Telomere Protection 225 GE52CH11_de_Lange ARI 26 October 2018 11:22 Figure 1 (Figure appears on preceding page) Shelterin structure and function. (a) Depiction of the six-subunit human shelterin complex associated with the double-stranded and single-stranded telomeric DNA. Note that mouse shelterin contains two POT1 proteins, POT1a and POT1b. TRF1 and TRF2 are shown as dimers, and the other shelterin subunits are each depicted once, although it is not known how many copies of each are present in the whole shelterin complex. (b) Depiction of shelterin complexes loaded onto telomeres in an open linear configuration and (c)in the t-loop configuration. Because TPP1 and POT1 are much less abundant than the other shelterin subunits, some complexes are depicted without TPP1 and POT1. (d ) Domain structure of the six human shelterin subunits and their protein and DNA interactions. Official gene names are given in parentheses. TRF2 is expressed as two functionally equivalent isoforms (L. Timashev & T. de Lange, unpublished data) either containing or lacking the N-terminal extension ( gray box). TIN2 has also been reported in a longer form. Splice variants of POT1 are not shown. The Myb/SANT domains of TRF1 and TRF2 are indicated with Myb. Reported and potential modification sites are indicated according to the key. (e) Summary of DDR pathways that are repressed by shelterin, the subunits dedicated to each pathway, and their proposed
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